My laboratory group is investigating the role of the RB tumor suppressor pathway in human cancer focusing primarily on the model of lung cancer since the clinical trials at the NCI-Navy Oncology Branch is a unique resource for patient material. In the past year, we have i) demonstrated that inactivation of the RB:p16 pathway occurs in 100% of lung cancer samples and have shown that inactivation of p16 can be reversed in vitro in a subset of lung cancer samples using the DNA demethylating agent, 5-aza2'deoxycytidine (AZAdc). We have characterized the kinetics of p16 induction with AZAdc and now plan to conduct a small clinical trial to test if p16 protein can also be induced in vivo; ii) We have studied the functional properties of several RB mutant alleles from families with the phenotype of low penetrance and have identified a unique type of "partial" mutant that characterizes these mutant proteins. We believe these mutants will address the molecular mechanisms that result in the phenotype of incomplete penetrance and will also be an invaluable resource in studying functional properties underlying the tumor suppressor activity of the RB product; iii) To study animal models of lung cancer, we have generated a colony of mice that are double heterozygous for null RB and p53 genes (common lesions in human small cell and non-small cell lung cancer) and have back-crossed these mice four times into the AJ mouse strain which is highly susceptible to lung tumor. We plan to study the pattern of tumor development and to analyze precursor cells for lung cancer; iv) We have characterized a novel pattern of post-translational modifications of the RB protein during apoptosis and are currently investigating the cellular localization of these specific cleavage products and whether they have a direct biological role in the process of programmed cell death.